1. Field of the Invention
This invention relates to a method for purifying an .alpha.-L-aspartyl-L-phenylalanine lower alkyl ester (hereinafter referred to as .alpha.-APE).
2. Description of the Prior Art
The .alpha.-APE, particularly .alpha.-L-aspartyl-L-phenylalaninemethyl ester (hereinafter referred to as .alpha.-APM), has a strong sucrose-like sweetness and thus has been drawing attention as novel sweetening.
Therefore, there have been proposed many methods for synthesizing the .alpha.-APE including, for example, a method of manufacturing it by directly bonding an L-phenylalaninemethyl ester and a strong acid addition salt of aspartic acid anhydride. Such a method, however, inevitably produces by-products such as a tripeptide ester which is formed by the coupling of the product .alpha.-APE with an additional molecule of aspartic acid, a tripeptide resulting from the tripeptide ester by the hydrolysis of an ester group, .alpha.-L-aspartyl-L-phenylalanine (hereinafter referred to as .alpha.-AP) resulting from the .alpha.-APE by the hydrolysis, a diketopiperazine derivative (hereinafter referred to as DKP), especially 3-benzyl-6-carboxymethyl-2,5-diketopiperazine, which results from the .alpha.-APE by the ring formation.
It is also known to obtain the .alpha.-APE by condensating an N-protected L-aspartic acid and an L-phenylalanine lower alkyl ester by an enzyme and then by removing the protection group. This method produces by-products to a lesser degree. However, even with this method employed, it is still hardly possible to substantially prevent .alpha.-AP and DKP production.
Conceivable methods for separating and purifying the .alpha.-APE would include a method of purifying the .alpha.-APE through a crystallization process. In this case, however, these by-products may tend to form mixed crystals because of their similarity with the .alpha.-APE in chemical structure. Therefore, these by-products still can be hardly removed by such a method.
In another known method for removing these by-products, an anion exchange resin of a specific type is employed. This method is disclosed in examined Japanese patent publication No. 35660/1977. In accordance with this method, these by-products are brought into contact with an anion exchange resin of an acetate or a formate type in an aqueous solvent to selectively remove them by adsorption. It is disclosed that it should not be possible to attain the object purification in the method if an other type of anion exchange resin than the organic acid addition type is employed because the impurity of the dipeptide ester material, i.e. an .alpha.-APE, would rather decrease if an anion exchange resin of such a type is used.
The present inventors conducted further studies on processes for purifying an .alpha.-APE using an anion exchange resin. As a result of the studies, they discovered that the purification of the .alpha.-APE using an anion exchange resin which had been believed to be impossible unless an anion exchange resin of an organic weak acid addition type was used could be quite effectively carried out by using a Cl-type anion exchange resin. This discovery has led to the present invention.